Expression of myelin genes in the developing chick retina

In submammalian animals including chicks, the retina contains oligodendrocytes (OLs), and axons in the optic fiber layer are wrapped with compact myelin within the retina; however, the expression of myelin genes in the chick retina has not been demonstrated yet. In the present study, we examined the expression of three myelin genes (proteolipid protein, PLP; myelin basic protein, MBP; cyclic nucleotide phosphodiesterase, CNP) and PLP in the developing chick retina, in comparison to the localization of Mueller cells. In situ hybridization demonstrated that all three myelin genes began to be expressed at E14 in the chick embryo retina. They are mostly restricted to the ganglion cell layer and the optic fiber layer, with a few exceptions in the inner nuclear layer where Mueller cells reside; however, PLP mRNA+ cells do not express glutamine synthetase, or vice versa. The present results elucidate that myelin genes are expressed only by OLs that are mostly localized in the innermost layer of the developing chick retina.     

The Msh-like homeobox genes define domains in the developing vertebrate eye.

The mouse Hox-7.1 gene has previously been shown to be related to the Drosophila Msh homeobox-containing gene. Here we report the isolation of a new member of this family which resides at an unlinked chromosomal location and has been designated Hox-8.1. Both Hox-7.1 and Hox-8.1 are expressed in the mouse embryo during the early stages of eye development in a distinct spatial and temporal relationship. Hox-8.1 is expressed in the surface ectoderm and in the optic vesicle before invagination occurs in regions corresponding to the prospective corneal epithelium and neural retina, respectively. Hox-7.1 is expressed after formation of the optic cup, marking the domain that will give rise to the ciliary body. The activity of these genes indicates that the inner layer of the optic cup is differentiated into three distinct compartments before overt cellular differentiation occurs. Our results suggest that these genes are involved in defining the region that gives rise to the inner layer of the optic cup and in patterning this tissue to define the iris, ciliary body and retina.     

Calretinin and calbindin in the retina of the developing chick

Summary Calretinin and calbindin-D28k are two calcium-binding proteins that are present in largely different sets of nerve cells in the central nervous system. Their appearance during development of the chick retina was studied by immunohistochemistry and Western blots. The patterns are mature one day before hatching. Each cell type acquires its characteristic calcium-binding protein several days after its differentiation has started, but in most cases before morphological maturation is complete. There is also an early phase of calbindin immunoreactivity in many immature amacrine cells, and of calretinin immunoreactivity in the presumptive photoreceptor layer, suggesting that these proteins may have distinct functions in differentiating cells.Abbreviations CR+ Immunoreactive for calretinin only - CB+ immunoreactive for calbindin only - CR+CB+ immunoreactive for both antisera - IPL inner plexiform layer - OPL outer plexiform layer     

Regulation of CLV3 expression by two homeobox genes in Arabidopsis

The ability of meristems to continuously produce new organs depends on the activity of their stem cell populations, which are located at the meristem tip. In Arabidopsis, the size of the stem cell domain is regulated by two antagonistic activities. The WUS (WUSCHEL) gene, encoding a homeodomain protein, promotes the formation and maintenance of stem cells. These stem cells express CLV3 (CLAVATA3), and signaling of CLV3 through the CLV1/CLV2 receptor complex restricts WUS activity. Homeostasis of the stem cell population may be achieved through feedback regulation, whereby changes in stem cell number result in corresponding changes in CLV3 expression levels, and adjustment of WUS expression via the CLV signal transduction pathway. We have analyzed whether expression of CLV3 is controlled by the activity of WUS or another homeobox gene, STM (SHOOT MERISTEMLESS), which is required for stem cell maintenance. We found that expression of CLV3 depends on WUS function only in the embryonic shoot meristem. At later developmental stages, WUS promotes the level of CLV3 expression, together with STM. Within a meristem, competence to respond to WUS activity by expressing CLV3 is restricted to the meristem apex.     

Spatiotemporal distribution of 1P1 antigen expression in the plexiform layers of developing chick retina

Changes in the distribution of 1P1-antigen in the developing chick retina have been examined by indriect immunofluorescence staining technique using the novel monoclonal antibody(MAb)1P1.Expression of the 1P1 antigen was found to be regulated in radial as well as in tangential dimension of the retina,being preferentially or exclusively located in the inner and outer plexiform layers of the neural retina depending on the stages of development ,With the onset of the formation of the inner plexiform layer 1P1 antigen becomes expressed in the retina.With progressing differentiation of the inner plexiform layer 1P1 immunofluorescence revealed 2 subbands at E9 and 6 subands at E18,At postnatal stages(after P3) immunoreactivity was reduced in an inside-outside sequence leading to the complete absence of the 1P1 antigen in adulthood.1P1 antigen expression in the outer plexiform layer was also subject to developmental regulation.The spation-temporal pattern of 1P1 antigen expression was correlated with the time course of histological differentation of chick retina,namely the synapse rich plexiform layers.Whether the 1P1 antigen was functionally involved in dendrite extension and synapse formation was discussed.     

Spatiotemporal distribution of 1P1 antigen expression in the plexiform layers of developing chick retina

Changes in the distribution of 1P1-antigen in the developing chick retina have been examined by indirect immunofiuorescence staining technique using the novel monoclonal antibody (MAb) 1P1. Expression of the 1P1 antigen was found to be regulated in radial as well as in tangential dimension of the retina, being preferentially or exclusively located in the inner and outer plexiform layers of the neural retina depending on the stages of development. With the onset of the formation of the inner plexiform layer 1P1 antigen becomes expressed in the retina. With progressing differentiation of the inner plexiform layer 1P1 immunofiuorescence revealed 2 subbands at E9 and 6 subbands at E18. At postnatal stages (after P3) immunoreactivity was reduced in an inside-outside sequence leading to the complete absence of the 1P1 antigen in adulthood. 1P1 antigen expression in the outer plexiform layer was also subject to developmental regulation. The spatio-temporal pattern of 1P1 antigen expression was correlated with the time course of histological differentiation of chick retina, namely the synapse rich plexiform layers. Whether the 1P1 antigen was functionally involved in dendrite extension and synapse formation was discussed.     

Expression of Frizzled genes in the developing chick eye

Frizzleds are transmembrane receptors that can transduce signals dependent upon binding of Wnts, a large family of secreted glycoproteins homologous to the Drosophila wingless (wg) gene product and critical for a wide variety of normal and pathological developmental processes. In the nervous system, Wnts and Frizzleds play an important role in anterior-posterior patterning, cell fate decisions, proliferation, and synaptogenesis. However, little is known about the role of Frizzled signaling in the developing eye. We isolated cDNAs for ten chick Frizzleds and analyzed the spatial and temporal expression patterns during eye development in the chick embryo. Frizzled-1 to -9 are specifically expressed in the eye at various stages of development and show a complex and partially overlapping pattern of expression.     

RCAS-RNAi: A loss-of-function method for the developing chick retina

Background  

The embryonic chick provides an excellent model system for studies of development. However, it has lacked an efficient loss-of-function method for studies of gene function.     

Comparative analysis of 3D expression patterns of transcription factor genes and digit fate maps in the developing chick wing

Hoxd13, Tbx2, Tbx3, Sall1 and Sall3 genes are candidates for encoding antero-posterior positional values in the developing chick wing and specifying digit identity. In order to build up a detailed profile of gene expression patterns in cell lineages that give rise to each of the digits over time, we compared 3 dimensional (3D) expression patterns of these genes during wing development and related them to digit fate maps. 3D gene expression data at stages 21, 24 and 27 spanning early bud to digital plate formation, captured from in situ hybridisation whole mounts using Optical Projection Tomography (OPT) were mapped to reference wing bud models. Grafts of wing bud tissue from GFP chicken embryos were used to fate map regions of the wing bud giving rise to each digit; 3D images of the grafts were captured using OPT and mapped on to the same models. Computational analysis of the combined computerised data revealed that Tbx2 and Tbx3 are expressed in digit 3 and 4 progenitors at all stages, consistent with encoding stable antero-posterior positional values established in the early bud; Hoxd13 and Sall1 expression is more dynamic, being associated with posterior digit 3 and 4 progenitors in the early bud but later becoming associated with anterior digit 2 progenitors in the digital plate. Sox9 expression in digit condensations lies within domains of digit progenitors defined by fate mapping; digit 3 condensations express Hoxd13 and Sall1, digit 4 condensations Hoxd13, Tbx3 and to a lesser extent Tbx2. Sall3 is only transiently expressed in digit 3 progenitors at stage 24 together with Sall1 and Hoxd13; then becomes excluded from the digital plate. These dynamic patterns of expression suggest that these genes may play different roles in digit identity either together or in combination at different stages including the digit condensation stage.     

Microenvironmental regulation of visual pigment expression in the chick retina

Visual pigment (VP) expression in the chick embryo retina was investigated in ovo, in dissociated and explant cultures, and in cDNAs from individual cells. While VP mRNA is not detectable by in situ hybridization until embryonic day (ED) 14-16 in ovo, analysis of VP expression by RT-PCR showed that VP messages are present in the retina as many as 7-10 days before they become detectable by in situ hybridization, and are also detected in other regions of the embryonic CNS. On the other hand, red opsin expression is markedly accelerated when cells are isolated from their intraocular microenvironment at ED 6, and placed in pigment epithelium-free dissociated or explant cultures. This acceleration occurs regardless of cell density, birth date, or serum presence in the medium, suggesting that many photoreceptors are already programmed to express red opsin on or before ED 6, and that microenvironmental inhibitory factors prevent implementation of this program until ED 14 in ovo. The selectivity of this phenomenon is suggested by the finding that other VPs are not observed by in situ hybridization in ED 6 cultures, although they are detectable in cultures of older retinas. Taken together, these findings suggest that red opsin expression may be constitutive for many developing photoreceptor cells in the chick.     

TGF-beta modulates programmed cell death in the retina of the developing chick embryo

Programmed cell death (PCD) is a key phenomenon in the regulation of cell number in multicellular organisms. We have shown that reduction of endogenous transforming growth factor beta (TGF-beta) prevents apoptotic PCD of neurons in the developing peripheral and central nervous system, suggesting that TGF-beta is an important mediator of ontogenetic neuron death. Previous studies suggested that there are other pro-apoptotic molecules, nerve growth factor (NGF) and brain-derived neurotrophic factor, that induce cell death in the nervous system. In the developing chick retina, NGF induces PCD by activation of the p75 receptor. We have studied the role of TGF-beta and its putative interdependence with NGF-mediated PCD in the chick retina. We found that TGF-beta is present in the developing chick retina during the period of PCD and is essentially required to regulate PCD of retinal cells. TGF-beta 2, TGF-beta 3 and the ligand-binding TGF-beta receptor can be detected immunocytochemically in the central retina, a region where apoptosis is most prominent during the early period of PCD. Application of a TGF-beta-neutralizing antibody to chick embryos in ovo resulted in a decrease in the number of TUNEL-positive cells and a reduction of free nucleosome levels. In terms of magnitude, reduction of PCD caused by the neutralization of endogenous TGF-beta was equivalent to that seen after anti-NGF application. Neutralization of both factors did not result in a further decrease in apoptosis, indicating that NGF and TGF-beta may act on the same cell population. Furthermore, neutralization of TGF-beta did not affect the expression of NGF or the p75-receptor. Our results suggest that TGF-beta and NGF are both required to regulate cell death in the chick retina in vivo.     

Differential expression of two msh-related homeobox genes Chox-7 and Chox-8 during chick limb development.

We have isolated two closely related cDNAs, Chox-7 and Chox-8, encoding homeodomain-containing proteins homologous to Drosophila msh. The Chox-7 and Chox-8 genes are chicken cognates of mouse Hox-7.1 and Hox-8.1, respectively. In situ hybridization using 3' regions of the cDNAs as probes revealed that the Chox-7 gene is highly expressed in the mesenchyme subjacent to the apical ectodermal ridge whereas Chox-8 expression is localized in the anterodistal mesenchymal region at early stages of limb formation, suggesting different roles during limb development. At later stages, both genes are expressed in the anterior and posterior mesenchymes and in the interdigital mesenchyme where programmed cell death occurs.